Ubon Ratchathani University

Improvement of drought Improvement of drought tolerant variety, IR57514 , for tolerant variety, IR57514 , for

grain aroma and cooking quality grain aroma and cooking quality using marker-assisted selectionusing marker-assisted selection

Ubon Ratchathani University

IR57514 is an elite drought tolerant and high yielding rice IR57514 is an elite drought tolerant and high yielding rice variety developed by IRRI which is well adapted to rainfed variety developed by IRRI which is well adapted to rainfed lowland in Mekong region where the areas are mostly limited lowland in Mekong region where the areas are mostly limited by abiotic stresses with unpredictable rainfall patterns. by abiotic stresses with unpredictable rainfall patterns.

Problem ?

BUT it was not

successfully adopted by farmers due to

unsatisfied of grain qualities.

Breeding IR57514 for grain aroma and good cooking quality traits will greatly enhance both quality and productivity of rice in the rainfed lowland areas in Mekong region.

IR57514 +aroma +

good cooking quality

The research was aimed to improve grain aroma and cooking quality of IR57514. Marker-assisted selection (MAS) in backcross breeding will be used to transfer favorable alleles of genes / QTL of grain aroma and good cooking qualities traits from KDML105 into genetic background of IR57514.

Objective

• The marker-assisted backcross breeding was conducted using IR57514 as the recipient parent. • KD571-77, the backcross introgression line of KDML105 carrying Xa21 for bacterial leaf blight resistance and Sub1A, Sub1B and Sub1C for submergence tolerance, was used as a donor parent for grain aroma and good cooking quality traits. • The backcross procedure for introgression of the target QTLs was conducted at Ubon Rice Research Center and Kasetsart University, Kamphangsean Campus, Nakorn Pathom.

Plant materialsPlant materials

Breeding scheme:

IR57514 x KD571-77Recipient parent Donor parent

F1 x IR57514

BC1F1

IR57514 x Selected BC1F1

BC2F1 (~6 line, ~27 seed)

~188 BC1F2MAS

IR57514 x selected BC1F2 (~18 plant)

90 BC2F1

MAS


BC3F1

11 selected BC3F1BC3F2 (1,640 plants)

436 Selected BC3F2 plants from 11 families

selected BC3F2 (33 plants)

MAS

Nov. 2007

July 2005

May, 2007

Jan. 2006

MAS

2,595 BC3F3

MAS

370 Selected BC3F3

May, 2008

MAS

Initial submergence tolerance testing (370 BC3F4 lines) Initial drought tolerance testing (319 BC3F4 lines)

173 selected lines

- YT in aerobic condition in KPS (173 BC3F4 lines)- YT in anaerobic condition in KPS (173 BC3F4 lines, stimulated flooding condition)

Yield trial(56 BC3F5 lines)

Trait validation (56 BC3F5 lines)- submergence tolerance- drought tolerance

Grain aroma and cooking quality testing

(56 BC3F5 lines)

August, 2008June, 2008

January, 2009

Preliminary grain aroma and cooking quality validation

56 selected lines


F1 x IR57514

BC1F1


BC2F1 (~6 line, ~27 seed)

~188 BC1F2MAS


90 BC2F1

MAS


BC3F1

11 selected BC3F1BC3F2 (1,640 plants)

436 Selected BC3F2 plants from 11 families


MAS

Nov. 2007

July 2005

May, 2007

Jan. 2006

MAS

2,595 BC3F3

MAS

370 Selected BC3F3

May, 2008

MAS

Initial submergence tolerance testing (370 BC3F4 lines) Initial drought tolerance testing (319 BC3F4 lines)

173 selected lines

- YT in aerobic condition in KPS (173 BC3F4 lines)- YT in anaerobic condition in KPS (173 BC3F4 lines, stimulated flooding condition)

Yield trial(56 BC3F5 lines)

Trait validation (56 BC3F5 lines)- submergence tolerance- drought tolerance

Grain aroma and cooking quality testing

(56 BC3F5 lines)

August, 2008June, 2008

January, 2009

Preliminary grain aroma and cooking quality validation

56 selected lines

• 352 BC3F1 were produced from the crosses between the selected 14 BC2F1 and IR57514.

•14 BC3F1 were selected based on

MAS:• BADH for grain aroma, • waxy for amylose content & gel consistency,• SNP 2341 for gelatinization temperature• PB7-8 for Xa21 bacterial leaf blight resistance

1st workshop (May 21 – 30, 2007)1st workshop (May 21 – 30, 2007)IR57514 x KD571-77

Recipient parent Donor parent

F1 x IR57514

BC1F1


~188 BC1F2

MAS


90 BC2F1

MAS


352 BC3F1

11 selected BC3F1

MAS

July 2005

1st workshop ( 21 -30 May, 2007)

Jan. 2006

MAS


F1 x IR57514

BC1F1


~188 BC1F2

MAS


90 BC2F1

MAS


352 BC3F1

11 selected BC3F1

MAS

July 2005

1st workshop ( 21 -30 May, 2007)

Jan. 2006

MAS

• 1640 BC3F2 plants were generated from 11 BC3F1

• The 436 BC3F2 progeny were selected from 1640 BC3F2 based on phenotype.

• 33 lines were selected using MAS:

• BADH for grain aroma, • waxy for amylose content & gel consistency,• SNP 2341 for gelatinization temperature• PB7-8 for Xa21 bacterial leaf blight resistance characters.

2nd workshop (Nov 10 – 24, 2007)2nd workshop (Nov 10 – 24, 2007)

BC3F2 (1,640 plants)

436 Selected BC3F2 plants


MAS

Preliminary quality validation

11 selected BC3F1


F1 x IR57514

BC1F1


~188 BC1F2

MAS


90 BC2F1

MAS


352 BC3F1

MAS

July 2005

Jan. 2006

MAS

May, 2007




MAS

Preliminary quality validation

11 selected BC3F1


F1 x IR57514

BC1F1


~188 BC1F2

MAS


90 BC2F1

MAS


352 BC3F1

MAS

July 2005

Jan. 2006

MAS

May, 2007

2nd workshop

• These 33 families of BC3F2 were selfing to generate 2,595 lines of BC3F3 progeny.

• Marker-assisted selection (MAS):

• BADH for aroma • waxy for amylose content and gel consistency• SNP2341 for gelatinization temperature and • PB 7-8 for Xa21 bacterial left blight resistance

• 370 BC3F3 progeny were selected based on genotype (260 lines) and based on phenotype (110 lines).

• The selected 370 BC3F3 progeny will be evaluated for the traits of interest comparing to the recipient variety, IR57514 .

3rd workshop (May 1 – 15, 2008)3rd workshop (May 1 – 15, 2008)




MASPreliminary quality

validation

11 selected BC3F1


BC1F1


90 BC2F1

MAS


352 BC3F1

MAS

Jan. 2006

MAS

May, 2007

2,595 BC3F3

MAS

370 Selected BC3F3




MASPreliminary quality

validation

11 selected BC3F1


BC1F1


90 BC2F1

MAS


352 BC3F1

MAS

Jan. 2006

MAS

May, 2007

2,595 BC3F3

MAS

370 Selected BC3F3

3rd workshop

Traits Validation• Grain quality: aroma, cooking quality

• BC3F2 • BC3F5

• Submergence tolerance• BC3F4• BC3F5

• Drought tolerance• BC3F4

4th workshop (May 1 – 15, 2008)4th workshop (May 1 – 15, 2008)

Grain aroma : sensory test

Amylose content : colorimetric method described by Julaino (1987)

Gelatinization temperature : Alkaline test (alkali spreading value ) as described by Little et al. (1958)

Gel consistency : described by Tan et al. (1999)

Traits Validation : Grain aroma and cooking quality in BC3F2 and BC3F5

Grain QualityGrain Quality

Traits Validation : Submergence tolerance (BC3F4 and BC3F5)Traits Validation : Submergence tolerance (BC3F4 and BC3F5)

Data recording ;

•Percent of plant survival (PPS)

• Percent of plant elongation (PPE)

• Leaf senescence

(Siangliw et al., 2003).

IR57514IR57514KDML105KDML105

FR13AFR13A

RD6RD6

Results 370 BC3F4 173 BC3F4

PPS

56 BC3F5 20 BC3F5

Location : Kasetsart University,

kampangsean campus,

Nakorn Pathom

PPS, LS, PPE

Percentage of plant survival

-5.000.005.00

10.0015.0020.0025.0030.0035.0040.0045.0050.0055.0060.0065.0070.0075.0080.0085.0090.0095.00

1 3 5 7 9 11 13 15 17 19 21 23 25 27 29 31 33 35 37 39 41 43 45 47 49 51 53 55 57 59

Lines

Pe

rce

nta

ge

of

pla

nt

su

rviv

al

RD

6K

DM

L105

IR57514

KD

571-7

7

Percentage of plant survival

-5.000.005.00

10.0015.0020.0025.0030.0035.0040.0045.0050.0055.0060.0065.0070.0075.0080.0085.0090.0095.00

1 3 5 7 9 11 13 15 17 19 21 23 25 27 29 31 33 35 37 39 41 43 45 47 49 51 53 55 57 59

Lines

Pe

rce

nta

ge

of

pla

nt

su

rviv

al

RD

6K

DM

L105

IR57514

IR57514

KD

571-7

7K

D571-7

7

Leaf greenness

0.00

10.00

20.00

30.00

40.00

50.00

1 4 7 10 13 16 19 22 25 28 31 34 37 40 43 46 49 52 55 58

Lines

SPAD

val

ue

IR57

514

KD57

1-7

7RD

6KD

ML1

05

Leaf greenness

0.00

10.00

20.00

30.00

40.00

50.00

1 4 7 10 13 16 19 22 25 28 31 34 37 40 43 46 49 52 55 58

Lines

SPAD

val

ue

IR57

514

KD57

1-7

7RD

6KD

ML1

05

Percent of plant elongation

0.00

20.00

40.00

60.00

80.00

100.00

1 3 5 7 9 11 13 15 17 19 21 23 25 27 29 31 33 35 37 39 41 43 45 47 49 51 53 55 57 59

Lines

perc

ent o

f pla

nt

elon

gatio

n KD57

1-7

7

RD6

KDM

L105

IR57

514

Percent of plant elongation

0.00

20.00

40.00

60.00

80.00

100.00

1 3 5 7 9 11 13 15 17 19 21 23 25 27 29 31 33 35 37 39 41 43 45 47 49 51 53 55 57 59

Lines

perc

ent o

f pla

nt

elon

gatio

n KD57

1-7

7

RD6

KDM

L105

IR57

514

IR57

514

Submergence tolerance validation in 56 BC3F5 lines. Submergence tolerance validation in 56 BC3F5 lines.

Validation : Drought tolerance (BC3F4) Validation : Drought tolerance (BC3F4)

Data recording

GY (Grain yield) PH (Plant height)

PN (Panicle number) TN (Tiller number)

FWD (Flowering date)

Experiment Design : Randomized Complete Block Design in CPA

Augmented Design in RGDU

Check varieties: IR57514 Tolerance check

KDML105, KD571-77 Susceptible check

Population size: 319 BC3F4 lines Chumpae Rice Research Center

173 BC3F4 lines Kasetsart University, Kampangsean campus

Grain aroma and cooking quality testing (BC3F5 progeny)Grain aroma and cooking quality testing (BC3F5 progeny)

Grain quality genotyping and validation

No.

# Aroma GT AC GC

Lines BADH Score SNP2341 Score Waxy Score Score

KD571-77 1 KD 1 5 1 16 73

IR57514 3 IR 3 2 3 33 102

1 BC3F5 # 14-34-1-16-1 1 KD* 1 4.9 1 19 86

2 BC3F5 # 22-43-1-10-19 1 KD 1 5.0 1 16 82

3 BC3F5 # 22-43-1-10-24 1 KD* 1 5.0 1 19 109

4 BC3F5 # 22-43-1-10-27 1 KD 1 4.9 1 20 106

5 BC3F5 # 22-43-1-10-32 1 KD 1 4.9 1 17 108

6 BC3F5 # 22-43-1-10-44 1 KD 1 4.9 1 19 146

7 BC3F5 # 22-43-1-42-25 1 KD 1 4.9 1 22 104

8 BC3F5 # 85-4-1-24-1 1 KD 1 5.0 1 20 123

9 BC3F5 # 85-4-1-24-12 1 KD 1 5.0 1 21 131

10 BC3F5 # 85-4-1-24-41 1 KD 1 5.0 1 21 106

Final Lines

Submergence tolerance and drought tolerance validationSubmergence tolerance and drought tolerance validation

No.

Lines

Submergence validation (BC3F5)

Drought validation (BC3F4)

Grain yield (kg rai-1) Sterility (%) Flowering date

PPS PPE LS Irrigated Drained Irrigated Drained Irrigated Drained

KD571-77 67.98 33.81 20.14 87.68 90.46 3.61 7.18 90 88

IR57514 73.39 27.21 31.99 140.82 121.40 4.37 4.93 103 98

1 BC3F5 # 14-34-1-16-1 79.29 24.93 33.04 101.40 103.47 1.92 0.67 80 81

2 BC3F5 # 22-43-1-10-19 73.61 21.50 27.81 - - - - - -

3 BC3F5 # 22-43-1-10-24 81.35 25.96 30.81 125.89 93.61 1.11 2.39 83 92

4 BC3F5 # 22-43-1-10-27 74.48 24.89 32.05 94.05 89.64 1.85 6.95 81 81

5 BC3F5 # 22-43-1-10-32 75.99 16.45 31.98 95.90 102.62 0.93 1.43 81 84

6 BC3F5 # 22-43-1-10-44 69.37 25.53 33.50 119.60 100.76 2.04 1.86 81 91

7 BC3F5 # 22-43-1-42-25 73.29 35.57 32.04 125.83 96.38 2.02 4.65 89 90

8 BC3F5 # 85-4-1-24-1 71.82 22.84 28.08 137.80 110.51 4.29 11.23 101 100

9 BC3F5 # 85-4-1-24-12 74.89 22.49 32.78 139.04 95.27 5.64 13.76 104 102

10 BC3F5 # 85-4-1-24-41 78.15 24.97 28.34 120.49 93.91 9.77 14.89 102 100

Mean 72.86 26.90 29.46 118.63 103.69 2.55 4.17 88.14 90.02

Prob. ** ** ** ** ** ** ** ** **

LSD0.05 2.36 1.67 0.63 1.68 1.48 0.17 0.30 0.68 0.64

Final Lines

Conclusion

We aimed to improve grain aroma and cooking quality of IR57514 using MAS.

10 selected BC3F5 were finally selected for grain aroma and cooking quality traits based on genotype (MAS) and these lines were validated for grain quality, submergence tolerance and drought tolerance.

These selected lines will be on trial both in station and on farm in rice production areas in Mekong region.

Marker assisted selection (MAS) technology was an excellent tool for rice breeding program.

MAS technology jointly with classical breeding will increase the effectiveness and precision of selection process.

MAS will be shortening the breeding program and also reduce the cost of breeding program.

Breeding IR57514 for grain aroma and good cooking quality traits will greatly enhance both quality and productivity of rice in the rainfed lowland areas in Mekong region.

Acknowledgments Acknowledgments

RRT H A I L A N Drice gene discovery UNIT

Rice Gene Discovery Unit Rice Gene Discovery Unit

(RGDU)(RGDU)

National Center for Genetic National Center for Genetic

Engineering and Engineering and

Biotechnology (BIOTEC)Biotechnology (BIOTEC)

Generation Challenge

Programe (GCP)

Documents

Ubon Ratchathani University